Regeneration

Question 1

How wound healing differs from regeneration

Answer 1

Regeneration is the renewal or restoration of a body part after injury
Wound healing is the process by which skin and tissue repairs itself after injury
unlike regeneration it has limited cell proliferation and an inflammatory response that activates fibroblasts

Question 2

Compensatory Regeneration

Answer 2

Differentiated cells divide to replace missing tissue but maintain their differentiated functions
ex: human liver
- remaining liver expands to compensate for lost parts

Question 3

Stem Cell based regeneration

Answer 3

New cells are routinely produced from stem cells to replace dying ones
ex: neoblasts in planarian allow it to reproduce asexually through binary fission

Question 4

Morphallaxis

Answer 4

The repatterning of existing tissues with little of no new cell growth. Cells may die, change shape, or be rearranged to restore the missing part. This result in a rescaling of the whole organism.
1. apoptosis eliminates some existing cells
2. transdifferentiation: existing cells change their identities to form new cell types required for regeneration
ex: hydra

Question 5

Epimorphosis

Answer 5

Lost tissue is replaced by the formation of a blastema (mass of undifferentiated cells)
a. the adult cells DEdifferentiate at the wound site, reverting them to an embryonic state
b. The dedifferentiated cells proliferate and differentiate to form the missing tissue
c. Cells may retain some memory of their previous identity (lineage restriction)

Question 6

Mechanism of Regeneration in Planaria
What happens following amputation of various sorts

Answer 6

Following amputation, planarians can regrow missing heads or tails. They use morphallaxis and stem cell regeneration. If a piece is too small, regeneration is abnormal.

Question 7

Mechanism of Regeneration in Planaria
cNeoblast vs specialized neoblasts

Answer 7

cNeoblasts are planarian pluripotent stem cells, they migrate to the wound site and regenerate tissues. They are scattered throughout the body except for the head and pharynx. They are constatnly replacing aging cells and responding to injury by migrating to the wound site.
Specialized neoblasts are descendants of cNeoblasts that have become committed to producing specific lineages of cells.

Question 8

Mechanism of Regeneration in Planaria
Use of radiation to study regeneration in planaria

Answer 8

Researchers have utilized radiation for “lose it” experiments that involve eliminating cNeoblasts. If all cNeoblasts are destroyed, regeneration is completely clocked and the planarian dies. A single surviving cNeoblast can repopulate the entire planarian.

Question 9

Mechanism of Regeneration in Planaria
How “lose it” experiments are done in planaria

Answer 9

RNAi
reverse genetics approach to achieve LOF gene mutation

Question 10

Mechanism of Regeneration in Planaria
Role of wnt signaling in planaria regeneration and how opposing gradients function in A/P

Answer 10

Low wnt - head
High wnt - tail
notum in the head is a wnt inhibitor, it cleaves plmitoleic acid off wnt which is required for wnt binding to frizzled.

Question 11

RNAi

Answer 11

1. introduce double stranded RNA corresponding to the target gene
   - in planarians this is done by feeding them bacteria that have been engineered to express the dsRNA
2. Inside the cell, dsRNA is processed by the ezyme Dicer (RNase enzyme that chops the long dsRNA into smaller fragments called siRNAs
3. The siRNAs then bind to a protein called RISC
4. RISC uses the siRNA as a guide to find mRNA, when RISC matches to mRNA, it cleaves it.
5. LOF for the gene since the cell cannot produce the protein

Question 12

Mechanism of Regeneration in Planaria
PCGs and muscle fiber organization that contribute to how cell’s position is remembered

Answer 12

Planarians possess a “morphological memory map” encoded by PCGs which guide the regeneration of missing parts with correct size and proportion. They are expressed throughout the planarian’s life, primarily in the muscle cells, providing a spatial coordinate system
The organization of muscle fibers, arranged in circular, diagonal, and longitudinal layers is linked to PCG expressiong.

Question 13

Mechanism of Regeneration in Planaria
Role of electrical potential in regeneration

Answer 13

Recent research suggeests that bioelectric patterns across tissues contribute to positional memory in planarians. Differences in membrane potential, influenced by ion channels exist along the A/P axis with the head being more depolarized.

Question 14

Key steps in salamander limb regeneration
Plasma clot

Answer 14

The first step in salamander limb regeneration, the plasma clot forms for wounding and healing. The clot also releases various signaling molecules which can acivate the inflammatory response

Question 15

Key steps in salamander limb regeneration
Wound epithelium

Answer 15

Epidermal cells from the limb stump cover the wound surface. As the migrating epidermal cells reach the wound site, they establish a new layer of skin called the wound epidermis. This thickened layer eventually transforms into the apical epidermal cap

Question 16

Key steps in salamander limb regeneration
Blastema formation

Answer 16

Blastema formation occurs due to epimorphic regeneration
The extracellular matrix beneath the wound epidermis is broken down, releasing cells that undergo dedifferentiation. These cells accumulate under the AEC to form the blastema

Question 17

Key steps in salamander limb regeneration
AEC and outgrowth

Answer 17

Signals from the AEC (thickened layer of epidermal cells) stimulates the growth and development of the blastema. The AEC secretes FGF8 which only happens in the presence of nerves. This is similar to the role of the AER in embryonic limb development. Without a sufficient number of nerve fibers, the AEC cannot exert growth promotion and regeneration is halted.

Question 18

How lineage analysis is done is axolotls and what it revealed about lineage restrictions during regeneration

Answer 18

Lineage analysis can be done by transplanting cells from a genetically modified donor axolotl expressing GFP into a host. Researchers can track the fate of the donor cells during regeneration. This research showed that there is lineage restriction meaning the cells in the blastema retain their lineage after dedifferentiation.

Question 19

Positional identity in axolotl limb regeneration
Rule of Distal transformation

Answer 19

A limb blastema will only generate the structures distal to the level at with the limb was amputated

Question 20

Positional identity in axolotl limb regeneration
Intercalary Regeneration

Answer 20

When tissues with different positional identities along the P/D axis are placed in contact with one another, this triggers the regeneration of intervening missing structures.

Question 21

In vivo evidence that blastemas “know” their position identity

Answer 21

By transplantation blastemas to different locations along the limb, researchers have observed that the regenerated structures strictly adhere to the positional identity of the blastema, not the site of transplantation.

Question 22

Molecular requirements for blastema proliferation
Response to ectopic RA

Answer 22

RA is a possible proximalizing factor. When applied to a regenerating salamander limb, RA can resent the blastema’s identity to a more proximal

Question 23

Molecular requirements for blastema proliferation
Role of nerves and AEC

Answer 23

Nerves signal Nrg 1 which is a requirement for blastema proliferation
Nerves support AEC formation and maintenance
AEC secretes FGF8 for blastema proliferation